A fully sealed intelligent nucleic acid extraction device with a microfluidic structure

Abstract

The present invention discloses a fully sealed intelligent nucleic acid extraction device with a microfluidic structure, comprising an extraction box, wherein the interior of the extraction box has a sample liquid chamber, a washing liquid chamber, an elution liquid chamber, a piston chamber and an extraction product liquid chamber which are vertically arranged and open at the bottom, wherein the sample liquid chamber has a sample addition port, and the sample addition port is provided with a sample addition sealing cover; the bottom surface of the extraction box is provided with a control valve; the piston chamber is provided with a piston rod, and during the lifting process of the piston rod, the liquid is driven to flow between the piston chamber and the sample liquid chamber, the washing liquid chamber, the elution liquid chamber or the extraction product liquid chamber connected to the piston chamber. The fully sealed intelligent nucleic acid extraction device of the present application has the advantages of being fully sealed, anti-pollution, easy to operate, low cost and capable of automation, and is particularly suitable for use in the fields of biomedical engineering or other sample testing instruments and equipment. It is particularly suitable for infectious biological samples to avoid the introduction of external contamination or harm to the outside world during the nucleic acid extraction process.

Description

Technical Field

[0001] The present invention relates to the technical field of nucleic acid extraction, and in particular to a fully sealed intelligent nucleic acid extraction device provided with a microfluidic structure. Background Art

[0002] Nucleic acid testing has become the main research method used in clinical molecular experiments. For example, the diagnosis and release of suspected patients infected with the new coronavirus require the support of nucleic acid test results. The premise of nucleic acid testing is to extract high-quality and high-purity nucleic acids from biological samples. Therefore, nucleic acid extraction and purification have become important steps in nucleic acid testing. Nucleic acid extraction methods are further divided into phenol chloride method, high salt precipitation method, silica gel centrifugal column chromatography method, ion exchange method, magnetic bead method, etc. Due to the advantages of no centrifugation, short operation time and stable method, the use of magnetic bead method to achieve nucleic acid extraction is increasing, and it has become one of the mainstream nucleic acid extraction methods.

[0003] The nucleic acid extraction process includes lysis, binding, washing, and elution. Currently, nucleic acid extraction is primarily performed manually in open laboratory environments, or each step is completed separately with the aid of instruments. This places high demands on the technicians, operating environment, and costs. Because biological samples are not sealed, they can easily be released into the external environment, causing contamination, especially for infectious biological samples. Furthermore, external contamination can be easily introduced during the nucleic acid extraction process. For example, the novel coronavirus infection poses a significant threat, and the detection of similar diseases requires a high degree of containment for nucleic acid extraction and testing.

[0004] Currently, there are very few instruments on the market that can complete fully automatic nucleic acid extraction in a single test kit, but the magnetic bead method is being used more and more. The magnetic bead method is used in the experimental devices described in "A genetic detection method, genetic detection kit and genetic detection equipment" with publication number CN 107151700 A and "A biological reaction device equipped with a microfluidic or nanofluidic structure" with publication number CN 109266518 A.

[0005] Currently, all fully sealed nucleic acid extraction kits on the market are used for extracting a single biological sample. For example, the experimental devices described in CN 111394221A "Enclosed Sample Processing Device" and CN 105695303 A "A Sample Reaction Device and Reaction Method" are circular and are used for extracting nucleic acids from a single biological sample. It is difficult to achieve the demand for simultaneous nucleic acid extraction from multiple biological samples in the same kit.

[0006] Currently available nucleic acid extraction devices with piston structures often feature a piston rod and flexible piston configuration, with the piston in direct contact with the liquid. For example, in the device described in CN 105695303 A, the liquid in the piston cavity is directly below the piston, resulting in a large gap between the piston head and the cavity. This results in a low liquid level when the eluent is minimal, affecting the nucleic acid extraction rate. Furthermore, flexible pistons made of materials other than special materials are hydrophilic, and if the piston comes into contact with the reagent, residual reagent liquid will inevitably remain on the piston.

[0007] Currently available piston-structured nucleic acid extraction devices using magnetic beads often store the magnetic beads below the piston cavity, lacking a dedicated cavity for storing the beads during the extraction process. For example, the device described in CN 111394221 A states that "a magnet captures the 'magnetic bead + DNA complex' on the inner wall of the piston cylinder 400." However, the magnetic beads are stored below the piston in the piston cavity. This approach makes it difficult to completely separate the bottom magnetic beads from the eluent, and the adsorption of magnetic beads on the inner wall can easily affect the piston's downward stroke, ultimately affecting the nucleic acid extraction rate. Summary of the Invention

[0008] In response to the above-mentioned deficiencies in the prior art, the present application provides a fully sealed intelligent nucleic acid extraction device with a microfluidic structure.

[0009] A fully sealed intelligent nucleic acid extraction device with a microfluidic structure, characterized by comprising an extraction box, wherein the extraction box has a vertically arranged sample liquid chamber with an open bottom surface, a washing liquid chamber, an eluent liquid chamber, a piston chamber, and an extraction product liquid chamber, wherein the sample liquid chamber has a sample addition port, and the sample addition port is provided with a sample addition sealing cover;

[0010] Before use, the sample liquid chamber, washing liquid chamber, and eluent chamber are pre-filled with lysis solution, washing solution, and eluent, respectively;

[0011] The bottom surface of the extraction box is provided with a control valve that can sequentially connect the piston chamber with the sample liquid chamber, the washing liquid chamber, the eluent liquid chamber, and the extraction product liquid chamber; a piston rod is provided in the piston chamber, and during the lifting and lowering process of the piston rod, liquid is driven to flow between the piston chamber and the sample liquid chamber, the washing liquid chamber, the eluent liquid chamber, or the extraction product liquid chamber connected to the piston chamber;

[0012] An air storage cavity is further provided inside the extraction box, and an exhaust channel is provided between the sample liquid cavity, the washing liquid cavity, the elution liquid cavity, the extraction product liquid cavity and the air storage cavity.

[0013] In order to facilitate the removal of samples after the extraction is completed, the extraction product liquid chamber has a sampling port, and the sampling port is provided with a sampling sealing cover. During the extraction process, the sampling sealing cover is in a covered state. After the extraction is completed, the product can be taken out by opening the sampling sealing cover. The structure is simple and convenient. The sealing cooperation between the sampling sealing cover and the sampling port can be sealed by inserting one end of a plug in an interference fit manner, that is, a plug portion is provided on the sampling sealing cover, and the plug portion is extended into the sampling port and is sealed by interference fit with the side wall of the sampling port. When the sampling sealing cover needs to be opened, the sampling sealing cover can be directly pulled out. Of course, the extraction product liquid chamber can also be designed as an assembled structure. After the extraction is completed, the extraction product liquid chamber can be directly removed from the entire extraction box and continue to be used as a container for storing samples.

[0014] In order to simplify the structure, the extraction product liquid chamber and the gas storage chamber are designed as one body.

[0015] The extraction box includes an extraction box body and a cover body that are sealed together. The cover body covers the top openings of the sample liquid cavity, the washing liquid cavity, the elution liquid cavity, the extraction product liquid cavity and the gas storage cavity. The exhaust channel is provided in the cover body. The peripheral edge of the cover body can extend toward the bottom side to form a fixed portion that cooperates with the outer periphery of the top of the extraction box body. The cover body and the extraction box body can be assembled and fixed by providing a snap-fit structure, or can also be fixed by providing a threaded fitting structure. Since the exhaust channel is provided inside the cover body, the extraction box is generally made of plastic. When the cover body is injection-molded, it is difficult to injection-mold the exhaust channel inside the cover body. Therefore, the cover body can be prepared as a split structure, and the top surface of the exhaust channel can be separately prepared as a cover sheet, and then the cover sheet and the remaining cover body parts are bonded together to form the cover body.

[0016] The control valve includes a translationally movable switching block having four flow channels that, during translation, connect the piston chamber to the sample, wash, eluate, or extract product chambers, respectively. Each extraction step is completed sequentially by switching to one of the four flow channels.

[0017] In one embodiment, the switching block includes an upper switching block and a lower switching block that cooperate with each other. The upper switching block has a guide hole that runs through the upper and lower parts, and the top surface of the lower switching block has a guide groove. The two guide holes are connected by connecting the guide grooves to form the guide channel. The guide grooves are 1 to 4, and the guide holes are four pairs. Each pair of guide holes cooperates with the guide groove to connect the piston chamber with one of the sample liquid chamber, the washing liquid chamber, the eluent chamber, or the extraction product liquid chamber. The four pairs of guide holes share or do not share the guide holes, and the four pairs of guide holes share or do not share the guide grooves. The relative movement between the upper switching block and the lower switching block allows the guide holes of the upper switching block and the guide grooves of the lower switching block to switch positions. When two guide holes correspond to the same guide groove, the two guide holes are connected through the same guide groove to form a guide channel. The guide holes used to form different guide channels can be shared with each other. Similarly, the guide grooves used to form different guide channels can also be shared with each other, as long as the requirements of forming four guide channels and switching between the guide channels when needed can be achieved.

[0018] The fully sealed intelligent nucleic acid extraction device also includes a base for mounting the extraction box. The base is provided with a mounting slot into which the bottom of the extraction box extends for secure installation. One end of the switching block extends into the mounting slot and is positioned below the extraction box. A side of the mounting slot is provided with a clearance for the other end of the switching block to extend. The sidewalls of the mounting slot and the outer surface of the extraction box are provided with mating buckle points and buckle holes.

[0019] The extraction box has a sealing membrane on its bottom surface, with avoidance holes formed in the membrane at locations corresponding to the bottom openings of the sample liquid chamber, wash liquid chamber, eluent liquid chamber, piston chamber, or extraction product liquid chamber. The top surface of the switching block is in contact with the sealing membrane and sealed against it. The sealing membrane enhances the sealing effect between each liquid chamber and the switching block, especially since the switching block needs to switch the connection status of each liquid chamber, which places high demands on the sealing effect.

[0020] The fully sealed intelligent nucleic acid extraction device of the present application can be used for magnetic bead extraction, and has the advantages of no centrifugation, short operation time, and stable method.

[0021] The piston rod is provided with a sealing ring at the end that extends into the piston cavity. A reduced diameter section is also provided at the end that extends into the piston cavity. A clearance space is formed between the reduced section and the sidewall of the piston cavity to allow the magnetic beads to escape. The sealing ring is located at the base of the reduced section. The base of the reduced section refers to the location of the reduced section away from the piston rod head. As the piston rod moves downward to expel liquid, the magnetic beads accumulate at the bottom of the piston cavity. The clearance space prevents the magnetic beads from being crushed by the piston rod and also prevents the piston rod from carrying away the accumulated magnetic beads upon contact, thereby reducing losses during nucleic acid extraction.

[0022] In order to facilitate the manipulation of magnetic beads during the separation and purification process and avoid damage to the magnetic beads due to the movement of the piston rod, a magnetic bead holding chamber is provided at the bottom of the side wall of the piston chamber, which sucks the magnetic beads used for adsorbing nucleic acids in the piston chamber under the action of an external magnetic force. In this way, the magnetic beads can be first controlled by magnetic force to be stored in the magnetic bead holding chamber, and then the piston rod can be manipulated to absorb and discharge related liquids, thereby reducing nucleic acid loss.

[0023] The extraction box has a flat structure. The sample liquid chamber, washing liquid chamber, elution liquid chamber, piston chamber and extraction product liquid chamber are arranged along the length direction of the extraction box. The magnetic bead holding chamber is located on one side of the bottom of the piston chamber side wall along the width direction of the extraction box.

[0024] The fully sealed intelligent nucleic acid extraction device comprises a plurality of extraction boxes, each of which is arranged side by side along the width direction of the extraction box. The fully sealed intelligent nucleic acid extraction device comprises 2 to 8 extraction boxes.

[0025] The fully sealed intelligent nucleic acid extraction device of the present application can efficiently extract biological samples. It only requires three reagents (lysis solution, washing solution, and eluent) and two movements (piston movement and switching block movement). It combines the lysis and binding processes in nucleic acid extraction into one, and simplifies other unnecessary operations such as drying, greatly simplifying the experimental process. The entire process can be completed in as little as 10 minutes.

[0026] The fully sealed intelligent nucleic acid extraction device of the present application can be used in conjunction with intelligent instruments. The entire process does not require manual operation or human supervision, making it more convenient for laboratory personnel to operate, thereby realizing the automation and intelligence of medical instruments in the fields of genetic testing and nucleic acid extraction.

[0027] The fully sealed intelligent nucleic acid extraction device of the present application adopts a microfluidic structure, and the nucleic acid extraction process can be performed even when the amount of reagent is extremely small.

[0028] The fully sealed intelligent nucleic acid extraction device of this application can extract nucleic acids more fully and the consistency of the extracted products is better. Using high-efficiency heat-resistant magnetic beads, the system can adsorb nucleic acids while lysing them. During the lysis process, all nucleic acid substances in the biological sample can be adsorbed with almost no residue. The piston-type magnetic bead liquid mixing method can make the magnetic beads spread throughout the liquid, with better mixing effect and easier adsorption of nucleic acid substances. The magnetic bead holding chamber in the piston cavity and the reduced diameter section on the lower side of the sealing ring lock the position of the magnetic beads, making them less likely to be contaminated on the piston or other locations, and the loss rate of nucleic acid substances is extremely small. The piston cooperates with the micro-flow guide channel to extract a quantitative eluent and then switch to the extraction product chamber for elution, eliminating the difference in eluent and ensuring consistent concentration of the extracted product.

[0029] The fully sealed intelligent nucleic acid extraction device, featuring a microfluidic structure, offers advantages such as complete enclosure, pollution prevention, ease of operation, low cost, and automation, making it particularly suitable for use in biomedical engineering and other areas of sample testing equipment. It is particularly suitable for use with infectious biological samples, avoiding the introduction of external contamination or potential harm during the nucleic acid extraction process. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 This is a structural schematic diagram of a fully sealed intelligent nucleic acid extraction device comprising only one extraction box according to the present invention.

[0031] Figure 2 This is a schematic diagram of the partial explosion structure of the fully sealed intelligent nucleic acid extraction device of the present invention which only includes one extraction box.

[0032] Figure 3 The figure is a schematic cross-sectional view of the fully sealed intelligent nucleic acid extraction device of the present invention which includes only one extraction box.

[0033] Figure 4 for Figure 3 A partial enlarged view.

[0034] Figure 5 This is a structural diagram of the switching block and the sealing membrane.

[0035] Figure 6 Schematic diagram of the structure of the switching block in the first embodiment.

[0036] Figure 7 Schematic diagram of the structure of the down switch block in the first embodiment.

[0037] Figure 8 FIG. 4 is a structural diagram of a switching block in the second embodiment.

[0038] Figure 9 Schematic diagram of the structure of the down switching block in the second embodiment.

[0039] Figure 10 Schematic diagram of the cross-sectional structure of the switching block in the second embodiment.

[0040] Figure 11 This is a structural diagram of the magnetic beads starting to be collected into the magnetic bead holding chamber under the action of an external magnetic force.

[0041] Figure 12 This is a structural diagram of the magnetic beads being collected into the magnetic bead holding cavity under the action of an external magnetic force.

[0042] Figure 13 Schematic diagram of the structure for canceling the external magnetic force.

[0043] Figure 14This is a structural diagram of the magnetic beads returning from the magnetic bead holding chamber to the piston chamber.

[0044] Figure 15 Schematic diagram of the structure of a fully sealed intelligent nucleic acid extraction device with two extraction boxes arranged side by side.

[0045] Figure 16 Schematic diagram of the structure of a fully sealed intelligent nucleic acid extraction device with four extraction boxes arranged side by side.

[0046] Figure 17 Schematic diagram of the structure of a fully sealed intelligent nucleic acid extraction device with 8 extraction boxes arranged side by side. DETAILED DESCRIPTION

[0047] like Figures 1 to 17 As shown, a fully sealed intelligent nucleic acid extraction device with a microfluidic structure can be used for nucleic acid extraction. The device offers advantages such as complete containment, pollution prevention, ease of operation, low cost, and automation. It is particularly suitable for use in biomedical engineering and other sample testing instrumentation and equipment. It is particularly suitable for infectious biological samples, preventing external contamination or harm during the nucleic acid extraction process.

[0048] like Figures 1 to 4 As shown, the fully sealed intelligent nucleic acid extraction device includes an extraction box 1, which has a vertically arranged sample liquid chamber 13, a washing liquid chamber 14, an eluent liquid chamber 15, a piston chamber 16, and an extraction product liquid chamber 17, each of which is open at the bottom. The sample liquid chamber 13 has a sample injection port 131, which is equipped with a sample injection sealing cap 132. Before use, the sample liquid chamber 13, the washing liquid chamber 14, and the eluent liquid chamber 15 are pre-filled with lysate, washing liquid, and eluent, respectively. The bottom surface of the extraction box 1 is provided with a control valve 2 that can sequentially connect the piston chamber 16 with the sample liquid chamber 13, the washing liquid chamber 14, the eluent liquid chamber 15, and the extraction product liquid chamber 17. A piston rod 161 is provided in the piston chamber 16. During the raising and lowering of the piston rod 161, the piston chamber 16 drives the liquid to flow between the piston chamber 16 and the sample liquid chamber 13, the washing liquid chamber 14, the eluent liquid chamber 15, or the extraction product liquid chamber 17 connected to the piston chamber 16. The bottoms of the sample liquid chamber 13 , the washing liquid chamber 14 , the elution liquid chamber 15 , the piston chamber 16 and the extraction product liquid chamber 17 may be designed to have an inner diameter that gradually decreases downward to facilitate liquid outflow and reduce liquid residue and dead volume.

[0049] Since the entire space inside the extraction box 1 is in a sealed state, in order to ensure that the air pressure in the local internal space does not change too much during operation and affect the flow of liquid, an air storage chamber is also provided inside the extraction box 1. An exhaust channel 121 is provided between the sample liquid chamber 13, the washing liquid chamber 14, the elution liquid chamber 15, and the extraction product liquid chamber 17. The air storage chamber serves as a buffer space for air pressure regulation. In the embodiment shown in the figure, in order to simplify the structure, the extraction product liquid chamber 17 and the air storage chamber are designed as one body.

[0050] In order to facilitate the removal of samples after the extraction is completed, the extraction product liquid chamber 17 has a sampling port 171, and the sampling port 171 is provided with a sampling sealing cover 172. During the extraction process, the sampling sealing cover 172 is in a covered state. After the extraction is completed, the sampling sealing cover 172 can be opened to remove the product. The structure is simple and convenient. The sealing cooperation between the sampling sealing cover 172 and the sampling port 171 can be sealed by inserting one end of a plug in an interference fit manner, that is, a plug portion is provided on the sampling sealing cover 172, and the plug portion is extended into the sampling port 171 to interfere with the side wall of the sampling port 171 to seal. When the sampling sealing cover 172 needs to be opened, the sampling sealing cover 172 can be directly pulled out. Of course, the extraction product liquid chamber 17 can also be designed as an assembled structure. After the extraction is completed, the extraction product liquid chamber 17 is directly removed from the entire extraction box 1 and continues to be used as a container for storing samples.

[0051] The extraction box 1 comprises an extraction box body 1 and a cover 12 that seal against each other. The cover 12 covers the top openings of the sample liquid chamber 13, the wash liquid chamber 14, the eluent liquid chamber 15, the extraction product liquid chamber 17, and the gas storage chamber. An exhaust passage 121 is provided within the cover 12. The peripheral edge of the cover 12 can extend toward the bottom side to form a fixed portion that mates with the top outer periphery of the extraction box body 11. The cover 12 and the extraction box body 11 can be assembled and fixed by providing a snap-fit structure or a threaded fitting structure. Of course, other assembly methods, such as gluing, ultrasonic welding, etc., can also be used. Since the exhaust channel 121 is arranged inside the cover body 12, the extraction box 1 is generally made of plastic. When the cover body 12 is injection molded, it is difficult to injection mold the exhaust channel 121 inside the cover body 12. Therefore, the cover body 12 can be prepared as a split structure, and the top surface of the exhaust channel 121 can be separately prepared as a cover piece 122, and then the cover piece 122 is bonded together with the remaining parts of the cover body 12 to form the cover body 12. Figure 3 : is a cross-sectional view of the fully sealed intelligent nucleic acid extraction device of the present application, wherein the middle position of the cover body 1 has an avoidance hole for the piston rod 161 to extend out, the exhaust channel 121 surrounds the avoidance hole position, and the exhaust channels 121 at various locations are interconnected.

[0052] like Figures 2 to 7As shown, control valve 2 includes a translatable switching block having four flow channels that, during translation, connect piston chamber 16 to sample liquid chamber 13, wash liquid chamber 14, eluent liquid chamber 15, or extracted product liquid chamber 17. The four flow channels connect piston chamber 16 to sample liquid chamber 13, wash liquid chamber 14, eluent liquid chamber 15, or extracted product liquid chamber 17, respectively. Each extraction step is completed sequentially by switching to one of the flow channels.

[0053] In one embodiment, the switching block includes an upper switching block 21 and a lower switching block 22 that cooperate with each other. The upper switching block 21 has a guide hole 23 that passes through the upper and lower parts, and the top surface of the lower switching block 22 has a guide groove 24. The two guide holes 23 are connected by connecting the guide grooves 24 to form a guide channel; there are 1 to 4 guide grooves 24, and there are four pairs of guide holes 23. Each pair of guide holes 23 cooperates with the guide grooves 24 to connect the piston cavity 16 with one of the sample liquid cavity 13, the washing liquid cavity 14, the elution liquid cavity 15 or the extraction product liquid cavity 17. The four pairs of guide holes 23 share or do not share the guide holes 23, and the four pairs of guide holes 23 share or do not share the guide grooves 24. The relative movement between the upper switching block 21 and the lower switching block 22 allows the positions of the guide holes 23 of the upper switching block 21 and the guide grooves 24 of the lower switching block 22 to be switched. When two guide holes 23 correspond to the same guide groove 24, the two guide holes 23 are connected through the same guide groove 24 to form a single guide channel. The guide holes 23 used to form different guide channels can be shared, and similarly, the guide grooves 24 used to form different guide channels can be shared, as long as the requirements of forming four guide channels and switching between the guide channels when needed can be achieved.

[0054] like Figures 4 to 7 In the embodiment shown in FIG, only four flow guide holes 23 arranged in a straight line are provided on the upper switching block 21, and only one flow guide groove 24 is provided on the top surface of the lower switching block 22. However, by translating the position of the switching block, the piston chamber 16 can be connected to the sample liquid chamber 13, the washing liquid chamber 14, the elution liquid chamber 15, the piston chamber 16, and the extraction product liquid chamber 17 through the cooperation and conduction with the bottom surface openings. For example, Figure 4The four guide holes 23 on the upper middle switching block 21 are respectively designated as the first, second, third, and fourth guide holes. When the first guide hole corresponds to the piston chamber 16, the fourth guide hole corresponds to the sample liquid chamber 13, and the piston chamber 16 and the sample liquid chamber 13 are connected via the guide groove 24 on the lower switching block 22. When the second guide hole corresponds to the piston chamber 16, the third guide hole corresponds to the wash liquid chamber 14, and the piston chamber 16 and the wash liquid chamber 14 are connected via the guide groove 24 on the lower switching block 22. When the third guide hole corresponds to the piston chamber 16, the second guide hole corresponds to the eluent chamber 15, and the piston chamber 16 and the eluent chamber 15 are connected via the guide groove 24 on the lower switching block 22. When the fourth guide hole corresponds to the piston chamber 16, the first guide hole corresponds to the extracted product liquid chamber 17, and the piston chamber 16 and the extracted product liquid chamber 17 are connected via the guide groove 24 on the lower switching block 22.

[0055] like Figures 8-10 This is another embodiment of the design of the guide holes 23' and the guide grooves 24'. The guide holes 23' are divided into two rows, one row with 2 holes and the other row with 4 holes. Correspondingly, the guide grooves 24' are also divided into two rows. There is a longer guide groove 24' corresponding to the two guide holes 23' in a row, and there are two shorter guide grooves 24' corresponding to the four guide holes 23' in a row. The piston chamber 16 and the sample liquid chamber 13 or the extraction product liquid chamber 17 located on both sides are connected through the longer guide groove 24' and the corresponding two guide holes 23'; through the two shorter guide grooves 24' and the corresponding four guide holes 23', a guide channel is formed between each shorter guide groove 24' and the corresponding two guide holes 23', one of which is used to connect the piston chamber 16 and the washing liquid chamber 14, and the other is used to connect the piston chamber 16 and the elution liquid chamber 15.

[0056] like Figures 1 to 3 As shown, the fully sealed intelligent nucleic acid extraction device of the present application also includes a base 3 for installing the extraction box 1. The base 3 is provided with a mounting groove 31. The bottom of the extraction box 1 extends into the mounting groove 31 for installation and fixation. One end of the switching block extends into the mounting groove 31 and is located below the extraction box 1. The side of the mounting groove 31 is provided with a avoidance opening 32 for the other end of the switching block to extend. The end of the switching block extending out of the avoidance opening 32 serves as an operating end and can cooperate with a corresponding intelligent operating instrument. The operating end can also be provided with a mounting hole 25 for cooperating with an intelligent operating instrument, or cooperate with an intelligent operating instrument in other forms. It only needs to be able to drive the switching block to move. Mutually cooperating buckle points 33 and buckle holes 18 are provided between the side wall of the mounting groove 31 and the outer side surface of the extraction box 1.

[0057] like Figures 3-5As shown, the bottom surface of the extraction cartridge 1 is provided with a sealing membrane 4, which has a receiving groove for accommodating the sealing membrane 4. The sealing membrane 4 is provided with avoidance holes 41 at positions corresponding to the bottom openings of the sample liquid chamber 13, the wash liquid chamber 14, the eluent liquid chamber 15, the piston chamber 16, or the extraction product liquid chamber 17. The top surface of the switching block is in contact with the sealing membrane 4, forming a seal. The sealing membrane 4 enhances the sealing effect between each liquid chamber and the switching block, especially since the switching block needs to switch the connection state of each liquid chamber, which places high demands on the sealing effect.

[0058] The fully sealed intelligent nucleic acid extraction device of the present application can be used for magnetic bead extraction, and has the advantages of no centrifugation, short operation time, and stable method.

[0059] like Figures 11-14 As shown, one end of the piston rod 161 extends out of the top surface of the cover body 12 and is used as an operating end. The piston rod 161 is provided with a sealing ring 162 at the end thereof extending into the piston cavity 16. The piston rod 162 is provided with a reduced diameter section 163 at the end thereof extending into the piston cavity 16. A clearance space 164 is formed between the reduced diameter section 163 and the side wall of the piston cavity 16 to avoid the magnetic beads 5. The sealing ring 162 is provided at the base of the reduced diameter section 163. The sealing ring 162 is provided at the base of the reduced diameter section 163, which refers to the portion of the reduced diameter section 163 away from the head of the piston rod 161 ( Figure 3 In this way, when the piston rod 161 moves downward to discharge the liquid, the magnetic beads 5 gather at the bottom of the piston chamber 16. Due to the provision of the avoidance space 164, the magnetic beads 5 can be prevented from being squeezed and crushed by the piston rod 161, and on the other hand, the loss of the extracted nucleic acid can be reduced.

[0060] In order to facilitate the manipulation of the magnetic beads 5 during the separation and purification process and avoid damage to the magnetic beads 5 due to the movement of the piston rod 161, a magnetic bead receiving chamber 165 is provided at the bottom of the side wall of the piston chamber 16, which sucks the magnetic beads 5 for adsorbing nucleic acids in the piston chamber 16 under the action of an external magnetic force. In this way, the magnetic beads 5 can be first controlled by magnetic force to be stored in the magnetic bead receiving chamber 165, and then the piston rod 161 can be manipulated to absorb and discharge related liquids, which can reduce nucleic acid loss.

[0061] The extraction box 1 is a flat structure, with the sample liquid chamber 13, the washing liquid chamber 14, the eluent chamber 15, the piston chamber 16, and the extraction product liquid chamber 17 arranged along the length of the extraction box 1. The magnetic bead receiving chamber 165 is located on the bottom side of the piston chamber 16 along the width of the extraction box 1. When the extraction box 1 is designed into this flat structure, the fully sealed intelligent nucleic acid extraction device of the present application can be designed into a multi-unit structure including multiple extraction boxes 1, each of which is arranged side by side along the width of the extraction box 1, for example Figures 15-17 There are 2-connected structures, 4-connected structures and 8-connected structures respectively, which can operate multiple samples at the same time and improve efficiency.

[0062] The fully sealed intelligent nucleic acid extraction device of the present application can efficiently extract biological samples, with only three reagents (lysis solution, washing solution, eluent) and two movements (piston movement, switching block movement), combining the lysis and binding processes in nucleic acid extraction into one, and simplifying other unnecessary operations such as drying, greatly simplifying the experimental process, and the whole process can be completed in as little as 10 minutes. The fully sealed intelligent nucleic acid extraction device of the present application can be used in conjunction with intelligent instruments. The whole process does not require manual operation or human supervision, which is more convenient for experimenters to operate, and realizes the automation and intelligence of medical instruments in the fields of genetic testing and nucleic acid extraction. The fully sealed intelligent nucleic acid extraction device of the present application adopts a microfluidic structure, and the nucleic acid extraction process can be carried out even when the amount of reagents is extremely small.

[0063] When in use, the entire nucleic acid extraction process can be divided into four steps: lysis, adsorption, washing, and elution. The lysis solution is pre-added in the sample liquid chamber 13. After the biological sample is added to the sample liquid chamber 13 through the sample addition port 131 on the top surface of the sample liquid chamber 13, it is immediately sealed with the sample addition sealing cover 132. After that, the entire extraction process is kept closed from the outside world; magnetic beads 5 are pre-added at the bottom of the piston chamber 16. The magnetic beads 5 use high-efficiency heat-resistant magnetic beads. The switching block in the control valve 2 is moved to the piston chamber 16 to connect with the sample liquid chamber 13, and the piston rod 161 is pulled up to make the liquid in the sample liquid chamber 13 be sucked into the piston chamber 16, lysing and adsorbing at the same time. During the lysis process, the nucleic acid of the biological sample is eluted. The acid substance can be completely adsorbed with almost no residue. During the lysis and adsorption process, the piston rod 161 can be pulled back and forth to help mix the magnetic beads 5 with the liquid, thereby helping lysis and adsorption. After the lysis and adsorption are completed, the magnetic control mechanism 6 is used to provide a magnetic field to accommodate the magnetic beads 5 in the magnetic bead receiving chamber 165, and the piston rod 161 is pushed downward to push the liquid back into the sample liquid chamber 13. The switching block in the control valve 2 is moved to the piston chamber 16 to connect with the washing liquid chamber 14. The washing liquid chamber 14 is pre-filled with washing liquid. The piston rod 161 is pulled up so that the liquid in the washing liquid chamber 14 is sucked into the piston chamber 16, and the magnetic control mechanism 6 is used again to provide a magnetic field to remove the magnetic beads 5 from the magnetic chamber 16. The beads 5 are released from the bead holding chamber 165 into the washing liquid. During the washing process, the piston rod 161 can be pulled back and forth to help mix the magnetic beads 5 with the liquid, thereby helping washing. After washing is completed, the magnetic control mechanism 6 provides a magnetic field to accommodate the magnetic beads 5 in the magnetic bead holding chamber 165, and the piston rod 161 is pushed downward to push the liquid back into the washing liquid chamber 14. The switching block in the control valve 2 is moved to connect the piston chamber 16 with the elution liquid chamber 15. The elution liquid chamber 15 is pre-filled with eluent, and the piston rod 161 is pulled up so that the liquid in the elution liquid chamber 15 is sucked into the piston chamber 16. The magnetic control mechanism 6 is used again to provide a magnetic field to remove the magnetic beads 5 from the magnetic bead holding chamber 165. The nucleic acid is released into the eluent. During the elution process, the piston rod 161 can be pulled back and forth to help mix the magnetic beads 5 with the liquid to help elution. After the elution is completed, the nucleic acid adsorbed on the magnetic beads is separated from the magnetic beads and enters the eluent. The magnetic control mechanism 6 provides a magnetic field to store the magnetic beads 5 in the magnetic bead holding chamber 165. Then, the switching block in the control valve 2 is moved to connect the piston chamber 16 with the extraction product liquid chamber 17. The piston rod 161 is pushed downward to push the eluent containing the nucleic acid into the extraction product liquid chamber 17 to complete the nucleic acid extraction step. After the extraction is completed, the sampling sealing cover 172 can be opened and the extraction product can be taken out through the sampling port 171. Of course, if some detection reagents are pre-added to the extraction product liquid chamber 17, the detection can be carried out directly after the extraction is completed without taking out the product. If so, the sampling port 171 corresponding to the extraction product liquid chamber 17 can also be directly not set.

Claims

1. A fully sealed intelligent nucleic acid extraction device with a microfluidic structure, characterized in that: The extraction box comprises an extraction box having a vertically arranged sample liquid chamber, a washing liquid chamber, an eluent liquid chamber, a piston chamber and an extraction product liquid chamber with an open bottom surface, wherein the sample liquid chamber has a sample addition port, and the sample addition port is provided with a sample addition sealing cover; Before use, the sample liquid chamber, washing liquid chamber, and eluent chamber are pre-filled with lysis solution, washing solution, and eluent, respectively; The bottom surface of the extraction box is provided with a control valve that can sequentially connect the piston chamber with the sample liquid chamber, the washing liquid chamber, the eluent liquid chamber, and the extraction product liquid chamber; a piston rod is provided in the piston chamber, and during the lifting and lowering process of the piston rod, liquid is driven to flow between the piston chamber and the sample liquid chamber, the washing liquid chamber, the eluent liquid chamber, or the extraction product liquid chamber connected to the piston chamber; The extraction box is further provided with an air storage cavity, and an exhaust channel is provided between the sample liquid cavity, the washing liquid cavity, the eluent liquid cavity, the extraction product liquid cavity and the air storage cavity; The control valve includes a translationally movable switching block having four flow guide channels for respectively connecting the piston chamber to the sample liquid chamber, the washing liquid chamber, the eluent liquid chamber, or the extraction product liquid chamber during translation; The extraction box is a flat structure, and the sample liquid chamber, washing liquid chamber, eluent liquid chamber, piston chamber and extraction product liquid chamber are arranged along the length direction of the extraction box; The switching block includes an upper switching block and a lower switching block that cooperate with each other. The upper switching block has a guide hole that passes through the upper and lower parts. The top surface of the lower switching block has a guide groove. The two guide holes are connected by the guide groove to form the guide channel. The piston rod is provided with a sealing ring at one end extending into the piston cavity; the piston rod is provided with a reduced diameter section at one end extending into the piston cavity, and a clearance space for the magnetic beads is formed between the reduced diameter section and the side wall of the piston cavity; the sealing ring is provided at the base of the reduced diameter section; The bottom of the side wall of the piston cavity is provided with a magnetic bead receiving cavity for sucking the magnetic beads for adsorbing nucleic acid in the piston cavity under the action of external magnetic force. The magnetic bead receiving cavity is located on one side of the bottom of the side wall of the piston cavity along the width direction of the extraction box.

2. The fully sealed intelligent nucleic acid extraction device according to claim 1, characterized in that: The extraction product liquid cavity has a sampling port, and the sampling port is provided with a sampling sealing cover; the extraction product liquid cavity and the gas storage cavity are designed as one body.

3. The fully sealed intelligent nucleic acid extraction device according to claim 1, characterized in that: The extraction box includes an extraction box body and a cover body that are sealed together. The cover body covers the top surface openings of the sample liquid chamber, the washing liquid chamber, the elution liquid chamber, the extraction product liquid chamber and the gas storage chamber. The exhaust channel is provided in the cover body.

4. The fully sealed intelligent nucleic acid extraction device according to claim 1, characterized in that: There are 1 to 4 guide grooves and four pairs of guide holes. Each pair of guide holes cooperates with the guide groove to connect the piston cavity and one of the sample liquid cavity, the washing liquid cavity, the elution liquid cavity or the extraction product liquid cavity. The four pairs of guide holes may share or not share the guide holes, and the four pairs of guide holes may share or not share the guide grooves.

5. The fully sealed intelligent nucleic acid extraction device according to claim 1, characterized in that: It also includes a base for mounting the extraction box, the base is provided with a mounting groove, and the bottom of the extraction box extends into the mounting groove for mounting and fixing; One end of the switching block extends into the installation slot and is located below the extraction box. A side of the installation slot is provided with an escape opening for the other end of the switching block to extend out.

6. The fully sealed intelligent nucleic acid extraction device according to claim 1, characterized in that: The bottom surface of the extraction box is provided with a sealing film, and the sealing film is provided with avoidance holes at positions corresponding to the bottom openings of the sample liquid chamber, washing liquid chamber, elution liquid chamber, piston chamber or extraction product liquid chamber; the top surface of the switching block is in contact with the sealing film and sealed.

7. The fully sealed intelligent nucleic acid extraction device according to claim 1, characterized in that: The utility model comprises a plurality of extraction boxes, and the extraction boxes are arranged side by side along the width direction of the extraction boxes.

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